|
| Status |
Public on Sep 16, 2008 |
| Title |
Input_DNA in HeLa S3 |
| Sample type |
SRA |
| |
|
| Source name |
Input DNA
|
| Organism |
Homo sapiens |
| Characteristics |
HeLa S3 cells (ATCC/Snyder Lab)
growth properties: Suspension Morphology: epithelial
cellular condition: Normal
|
| Growth protocol |
HeLaS3 cells were grown in Minimum Essential Medium modified for Suspension (SMEM, Invitrogen), supplemented with glutamine, 10% FBS, and antibiotics (penicillin-streptomycin) at 37C in 5% CO2, to a density of 5 x10^5 cells/ml.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Purified ChIP and Input DNA samples from 2 x 10^8 cells were size-selected on agarose gels for fragments 120 to 350 bp prior to library preparation. RNA polymerase II ChIP DNA samples were prepared according to the same protocol as used for the STAT1 samples except that the cells were not stimulated prior to formaldehyde fixation. RNA Pol II-DNA complexes were isolated with the mouse monoclonal antibody 8WG16 (Covance). Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
Input DNA
|
| Data processing |
Enriched peak regions corresponding to transcription factor binding sites were identified by first extending all the uniquely mapped tags reads (mapped against NCBI36) to the averaged sequenced DNA fragment size ~200bps and then accumulated into fragment density signal maps. Peaks in these maps are identified by comparison to a computer simulated null model for each Mb of each chromosome (accounting for the variability in mappability of sequence tags across the human genome. For potential target regions the number of sequenced tags is compared against the normalized number of sequence tags overlapping the region from the matching Input DNA reference sample. Fold enrichment ratios are computed from the ratio of these two sets of counts for each regions as well as p-value using the cumulative distribution function for the binomial distribution. P-value are corrected for multiple hypothesis testing using a Benjamini-Hochberg correction factor and a false discovery rate threshold of 0.05 is imposed.
|
| |
|
| Submission date |
Sep 15, 2008 |
| Last update date |
May 15, 2019 |
| Contact name |
Flora Vaccarino |
| Organization name |
Yale University
|
| Department |
Child Study Center
|
| Lab |
Vaccarino
|
| Street address |
230 South Frontage Road
|
| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06520 |
| Country |
USA |
| |
|
| Platform ID |
GPL9115 |
| Series (2) |
| GSE12781 |
PolII transcription factor in Human HeLa S3 |
| GSE12783 |
Genome-wide maps of transcription factor binding using ChIP-Seq for Human ENCODE |
|
| Relations |
| SRA |
SRX003802 |
| BioSample |
SAMN02195758 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneA_FC6144_082907_s_5.txt.gz |
32.1 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneB_FC12033_091907_s_4.txt.gz |
88.2 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneC_FC12033_091907_s_5.txt.gz |
77.5 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneD_FC12033_091907_s_6.txt.gz |
86.1 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneE_FC12033_091907_s_7.txt.gz |
74.1 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneF_FC12044_091407_s_8.txt.gz |
69.0 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneG_FC12060_090907b_s_7.txt.gz |
49.6 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneH_FC12170_102207_s_1.txt.gz |
123.8 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneI_FC12170_102207_s_2.txt.gz |
129.1 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneJ_FC12170_102207_s_3.txt.gz |
110.0 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneK_FC12170_102207_s_4.txt.gz |
136.4 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneL_FC12187_102707_s_1.txt.gz |
119.1 Mb |
(ftp)(http) |
TXT |
| GSM320735_HeLa_Input_DNA_eland_results_rep1_laneM_FC12187_102707_s_2.txt.gz |
118.5 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
